A research program is proposed on the induction and consequences of protein synthesis errors in eukaryotic cells. This program is based on findings made in our laboratory, concerning the stimulation of misreading by paromomycin, and certain other structurally related members of the aminoglycoside antibiotics. The ultimate goal is to identify the genes controlling the accuracy of ribosomes in eukaryotic cells and to describe the action of these genes in molecular detail. We have isolated several mutants of the ciliated protozoan, Tetrahymena, which are resistant to paromomycin; also, we have identified a mutant of the yeast, Saccharomyces cerevisiae, which is super-sensitive to the drug. In these organisms, we plan to conduct genetic analysis of these mutants, and to isolate others with altered responses to misreading antibiotics. We will also conduct a biochemical analysis to determine if the mutations alter the structure of the ribosomes and the accuracy of protein synthesis. We have shown that paromomycin suppresses nonsense mutations in yeast; we plan to isolate mutants of Tetrahymena with suppressible defects, to extend the potential of genetic analysis in the organism. We also plan biochemical studies on the binding of radiolabeled antibiotics to ribosomes of normal and mutant organisms. Finally, we have observed that certain stresses, including misreading antibiotics and heat, dramatically induce the synthesis of some specific polypeptides in Tetrahymena. We will attempt to correlate this induction with changes in enzymatic activities, particularly in the pathways of gluconeogenesis, amino acid catabolism, and protein degradation.